Systems pharmacology-based mechanism exploration of Acanthopanax senticosusin for Alzheimer's disease using UPLC-Q-TOF-MS, network analysis, and experimental validation.

BACKGROUND: Alzheimer's disease (AD) is a neurodegenerative disease, characterized by progressive cognitive dysfunction and memory loss. However, the disease-modifying treatments for AD are still lacking. Traditional Chinese herbs, have shown their potentials as novel treatments for complex diseases, such as AD. PURPOSE: This study was aimed at investigating the mechanism of action (MOA) of Acanthopanax senticosusin (AS) for treatment of AD. METHODS: In this study, we firstly identified the chemical constituents in Acanthopanax senticosusin (AS) utilizing ultra-high performance liquid chromatography coupled with Q-TOF-mass spectrometry (UPLC-Q-TOF-MS), and next built the drug-target network of these compounds. We also performed the systems pharmacology-based analysis to preliminary explore the MOA of AS against AD. Moreover, we applied the network proximity approach to identify the potential anti-AD components in AS. Finally, experimental validations, including animal behavior test, ELISA and TUNEL staining, were conducted to verify our systems pharmacology-based analysis. RESULTS: 60 chemical constituents in AS were identified via the UPLC-Q-TOF-MS approach. The systems pharmacology-based analysis indicated that AS might exert its therapeutic effects on AD via acetylcholinesterase and apoptosis signaling pathway. To explore the material basis of AS against AD, we further identified 15 potential anti-AD components in AS. Consistently, in vivo experiments demonstrated that AS could protect cholinergic nervous system damage and decrease neuronal apoptosis caused by scopolamine. CONCLUSION: Overall, this study applied systems pharmacology approach, UPLC-Q-TOF-MS, network analysis, and experimental validation to decipher the potential molecular mechanism of AS against AD.

Covalent Modification of Proteins by Osthole Reactive Metabolites using Proteomic Approaches.

BACKGROUND: Osthole (OST) is a bioactive natural coumarin derived from the plant Cnidium monnieri (L.) Cusson fruit (She Chuang Zi), which has various pharmacological and biological activities. OST contains an α,ß- unsaturated lactone, which is an electrophilic group that tends to be metabolized into reactive metabolites (RMs). Then, RMs are able to covalently modify nucleophilic amino acid (AA) residues of target proteins. However, few researchers considered the contribution of the covalent modification induced by OST or its metabolites. OBJECTIVE: This study aims to investigate the metabolic profile and the metabolites-protein modification of OST. METHODS: The metabolites of OST were qualitatively identified using UHPLC-Q-TOF-MS. The RMs modification patterns and potentially modified AA residues were confirmed by UHPLC-Q-TOF-MS using rat liver microsomes (RLMs) and model AAs. Finally, the modified peptides derived from high-abundance microsomal peptides were separated via nano-LC-Orbitrap-MS, and then RM-modified proteins were identified using a proteome discoverer. RESULTS: In the presence of RLMs, OST could rapidly be metabolized within 1 h and hardly identified at 4 h. We detected 10 OST metabolites, 13 OST metabolites-NAC (N-acetyl cysteine) adducts, 3 NAL (N-acetyl lysine) adducts, and 11 GSH (glutathione) adducts. Furthermore, 16 RM-modified protein targets were identified, many of which are included in the essential biological processes of OST's anti-Alzheimer's disease (AD) and anti-tumor. CONCLUSION: This study provides a novel perspective on the molecular mechanism of OST's pharmacological activities, as well as identifies potential targets for further development and application of OST and other Natural products (NPs).

Ferulic acid prevents Diosbulbin B-induced liver injury by inhibiting covalent modifications on proteins.

Diosbulbin B (DIOB) has been reported to cause serious liver injury. However, in traditional medicine, DIOB-containing herbs are highly safe in combination with ferulic acid (FA)-containing herbs, suggesting potential neutralizing effect of FA on the toxicity of DIOB. DIOB can be metabolized to generate reactive metabolites (RMs), which can covalently bind to proteins and lead to hepatoxicity. In the present study, the quantitative method was firstly established for investigating the correlation between DIOB RM-protein adducts (DRPAs) and hepatotoxicity. Then, we estimated the detoxication effect of FA in combination with DIOB and revealed the underlying mechanism. Our data indicated that the content of DRPAs positively correlate with the severity of hepatotoxicity. Meanwhile, FA is able to reduce the metabolic rate of DIOB in vitro. Moreover, FA suppressed the production of DRPAs and decreased the serum alanine/aspartate aminotransferase (ALT/AST) levels elevated by DIOB in vivo. Thus, FA can ameliorate DIOB-induced liver injury through reducing the production of DRPAs.

Gallic acid enhances anti-lymphoma function of anti-CD19 CAR-T cells in vitro and in vivo.

Chimeric antigen receptor T (CAR-T) cell targeting CD19 antigen has achieved exhilarative clinical efficacy in B-cell malignancies. However, challenges still remain for the currently approved anti-CD19 CAR-T therapies, including high recurrence rates, side effects and resistance. Herein, we aim to explore combinatorial therapy by use of anti-CD19 CAR-T immunotherapy and gallic acid (GA, an immunomodulatory natural product) for improving treatment efficacy. We assessed the combinatorial effect of anti-CD19 CAR-T immunotherapy with GA in cell models and a tumor-bearing mice model. Then, the underlying mechanism of GA on CAR-T cells were investigated by integrating network pharmacology, RNA-seq analysis and experimental validation. Furthermore, the potential direct targets of GA on CAR-T cells were explored by integrating molecular docking analysis with surface plasmon resonance (SPR) assay. The results showed that GA significantly enhanced the anti-tumor effects, cytokine production as well as the expansion of anti-CD19 CAR-T cells, which may be mainly through the activation of IL4/JAK3-STAT3 signaling pathway. Furthermore, GA may directly target and activate STAT3, which may, at least in part, contribute to STAT3 activation. Overall, the findings reported here suggested that the combination of anti-CD19 CAR-T immunotherapy with GA would be a promising approach to increase the anti-lymphoma efficacy.

Combination of chemical profiling and network pharmacology analysis to investigate the potential mechanism of Li-Zhong-Xiao-Pi granules in the treatment of gastric precancerous lesions.

Li-Zhong-Xiao-Pi granules (LZXP) are effective for treating gastric precancerous lesions (GPL) in traditional Chinese medicine. However, the active compounds of LZXP and their potential therapeutic mechanism in GPL remained unclarified. The purpose of this study is to investigate the chemical composition and potential targets of LZXP. Based on the accurate masses, ion fragments, and literature data, a total of 128 compounds were identified in the LZXP sample using ultra-performance liquid chromatography and quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) in both positive and negative ion modes, and 28 of these compounds were exactly determined by comparison with authentic reference standards. Meanwhile, 11 typical components were quantified via UPLC during a 24 min period. The linearity, accuracy, stability and recovery of the method were all proven. Through the network pharmacological analysis, six chemicals (quercetin, 4'-hydroxywogonin, sinensetin, 5, 7, 8, 3', 4'-pentamethoxyflavanone, 8-gingerdione and quercetin) were identified as the active ingredients, and five LZXP targets (AKT1, CYP1B1, PTGS2, MMP9 and EGFR) were found to be the crucial molecules in the treatment of GPL. This study provides a systematic and applicable method for the rapid screening and identification of the chemical constituents from LZXP, and an effective understanding the mechanism of LZXP in the treatment of GPL.

Construction and functional identification of CD19-CAR vector based on PiggyBac transposable system / 中国肿瘤生物治疗杂志

@#[摘 要] 目的：开发基于PiggyBac（PB）转座系统的电转染CAR-T细胞制备方法并鉴定其体外抗肿瘤功能。方法：采用健康人外周血单个核细胞（PBMC）制备T细胞，通过分子克隆技术将CD19基因克隆到PB质粒（转座子）中后经电转染法将转座子和转座酶质粒导入激活的T细胞中，并测定其转染效率，最后运用流式细胞术及荧光素酶发光实验评估其对人Burkitt's淋巴瘤Raji细胞的杀伤能力。结果：电转染制备的CD19 CAR-T细胞转染效率较高（>60%），呈剂量依赖性，且CAR-T细胞相对于Pan-T细胞对Raji细胞杀伤能力显著（P<0.05）。结论：开发的PB转座系统的电转染方法可行，在体外对肿瘤细胞具有显著的杀伤能力，具备临床运用于CD19 CAR-T细胞制备的潜力。

Evolving Roles of Natural Terpenoids From Traditional Chinese Medicine in the Treatment of Osteoporosis.

Osteoporosis (OP) is a systemic metabolic skeletal disease which can lead to reduction in bone mass and increased risk of bone fracture due to the microstructural degradation. Traditional Chinese medicine (TCM) has been applied in the prevention and treatment of osteoporosis for a long time. Terpenoids, a class of natural products that are rich in TCM, have been widely studied for their therapeutic efficacy on bone resorption, osteogenesis, and concomitant inflammation. Terpenoids can be classified in four categories by structures, monoterpenoids, sesquiterpenoids, diterpenoids, and triterpenoids. In this review, we comprehensively summarize all the currently known TCM-derived terpenoids in the treatment of OP. In addition, we discuss the possible mechanistic-of-actions of all four category terpenoids in anti-OP and assess their therapeutic potential for OP treatment.

Co-Expression of miR155 or LSD1 shRNA Increases the Anti-Tumor Functions of CD19 CAR-T Cells.

Chimeric antigen receptor (CAR) T cells targeting CD19 antigen have produced remarkable clinical outcomes for cancer patients. However, identifying measures to enhance effector function remains one of the most challenging issues in CD19-targeted immunotherapy. Here, we report a novel approach in which a microRNA (miRNA) or short-hairpin RNA (shRNA) cassette was integrated into CAR-expressing retroviral vectors. Using this system, we generated anti-CD19 CAR-T cells co-expressing miR155 or LSD1 shRNA and found that anti-CD19 CAR-T cells with miR155 upregulation or LSD1 downregulation exhibited increased anti-tumor functions in vitro and in vivo. Transcriptional profiling analysis by RNA sequencing revealed the targets of miR155 and LSD1 in anti-CD19 CAR-T cells. Our experiments indicated that introduction of miRNA or shRNA expression into anti-CD19 CAR T-cells might be an effective strategy to improve the anti-tumor effects of CAR-T cell therapy.

Multimodal intraoperative monitoring during surgical correction of scoliosis to avoid neurologic damage.

The purpose of this study was to evaluate the application of multimodal intraoperative monitoring (MIOM) system in patients with congenital scoliosis (CS) and adolescent idiopathic scoliosis (AIS).Twelve patients who underwent posterior surgical correction of scoliosis for CS and AIS from June 2014 to July 2018 were enrolled in this study. During the operation, we monitored the functional status of the spinal cord by MIOM. An abnormal somatosensory evoked potential was defined as a prolonged latency of more than 10% or a peak-to-peak amplitude decline of more than 50% when compared to baseline. An abnormal transcranial motor evoked potential (TcMEP) was defined as a TcMEP amplitude decrease of more than 50%. A normal triggered electromyography response, which presented with the absence of an electrical response on stimulation at 8.2 mA, indicated that the pedicle screw was not in contact with the spinal cord or nerve root.A total of 12 patients underwent MIOM surgery, of which 9 patients with negative MIOM had no significant deterioration of neurological function postoperatively, and exhibited satisfactory surgical correction of scoliosis during follow-ups. However, the remaining 3 patients suffered from MIOM events, 2 patients had normal neurological function, and 1 patient had deteriorated neurological function postoperatively.Using MIOM in CS and AIS surgery could promptly detect iatrogenic neurological injury at the early stage. Therefore, rapid response by appropriate intraoperative interventions can be taken to minimize the injury. Besides, stable MIOM recordings encourage surgeons to correct scoliosis even when the Cobb angle of scoliosis was extremely large.

A screw-view model of navigation aid retrograde transpubic screw fixation for anterior pelvic ring fracture: A case report with 28 months follow-up and technical note.

RATIONALE: The purpose of this study was to evaluate the accuracy of a retrograde transpubic screw fixation assisted by a screw-view model of navigation in treating a pelvic fracture. PATIENT CONCERNS: A 30-year-old female patient injured in a motor vehicle accident, displayed symptoms characterized by swelling and pain of the pudendum. DIAGNOSES: The patient was diagnosed with a fracture of the pubic ramus. INTERVENTIONS: We used a screw-view model of navigation to assist our retrograde transpubic screw fixation in this patient. OUTCOMES: In total, 2 screws were inserted into the bilateral pubic ramus and both were excellently positioned. It took 7.4 minutes to design the screws, 8.1 minutes to implant the guidewire, and 39.3 minutes to place the screws. Intraoperative blood loss amounted to 21 mL and the total fluoroscopic time was 3.8 minutes. No clinical complications, such as neurologic, vascular, or urologic injury, infection, screw loosening, or loss of reduction, were found after the operation. Follow-up lasted 28 months. LESSONS: The outcome of our study suggests that the screw-view model of navigation maximizes the retrograde transpubic screw insertion accuracy in the treatment of a pubic ramus fracture, which is made efficient by pain relief and early out-of-bed mobilization. Our suggestion is, therefore, that the relative position between the pubic ramus and the patient tracker must be static to ensure the accuracy of the entire system throughout the operation.

Accuracy and safety of percutaneous periacetabular screw insertion using screw view model of navigation in acetabular fracture: A case report.

RATIONALE: The purpose of this study was to estimate the efficacy and safety of percutaneous periacetabular screw (PPS) insertion assisted by screw view model of navigation (SVMN) to treat fracture of acetabulum. PATIENT CONCERNS: A 61-year-old male patient was injured in a motorcycle accident, which caused pain, swelling, deformity and limited mobility on his right hip. DIAGNOSES: He was diagnosed with fracture of acetabulum. INTERVENTIONS: We used PPS insertion assisted by SVMN to treat fracture of acetabulum in this patient. OUTCOMES: The follow up lasted 24 months. Totally 2 screws were inserted into anterior and posterior column of acetabulum respectively and both of them displayed grade 0. Compared with the preoperative gap and step of fracture displacement, the postoperative ones were significantly reduced. It took 11.7 minutes for designing the screws, 6.7 minutes for implanting the guide wire, and 45.5 minutes for placing the screws. Intraoperative blood loss was 29 mL and total fluoroscopic time was 4.1 minutes. No clinical complications such as nerve vascular injury, infection and screw loosening were found after the operation. LESSONS: The study indicated that SVMN is favorable to the PPS insertion for acetabular fracture. Our lesson is that the relative position between the acetabular and the patient tracker must be static to ensure the accuracy of the entire system throughout the operation.

Multimodal intraoperative monitoring during reduction of spine burst fracture and dislocation prevents neurologic injury.

This study aims to evaluate the application of multimodal intraoperative monitoring (MIOM) in surgical treatment for spine burst fracture and dislocation (SBFD) patients.Eleven patients who underwent posterior reduction and instrumentation (PRI) for SBFD from June 2014 to July 2016 were included into the study. The function of the spinal cord was monitored by MIOM. The muscle strength of the lower extremities and American Spinal Injury Association (ASIA) scores were, respectively, evaluated (before surgery, and at 1, 3, 6, and 12 months after surgery). Furthermore, the extent of reduction was also assessed.Muscle strength recovery, ASIA score changes, and the extent of reduction were correlated with MIOM results. Among the 11 patients who received surgery under MIOM, 8 patients with negative MIOM results during the operation did not demonstrate neurological deterioration postoperatively and exhibited improvements in ASIA scores during follow-ups. However, among the 3 patients who encountered MIOM events (case 4, 7, and 8), 2 patients avoided nerve lesion and 1 patient suffered from neurologic deterioration postoperatively.The application of MIOM technology during PRI surgery may detect spinal cord impairment at the early stage, and operative schemes can be modified before permanent nerve compromise is triggered by surgical manipulation.

Identification of key genes associated with rheumatoid arthritis with bioinformatics approach.

We aimed to identify key genes associated with rheumatoid arthritis (RA).The microarray datasets of GSE1919, GSE12021, and GSE21959 (35 RA samples and 32 normal controls) were downloaded from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) in RA samples were identified using the t test in limma package. Functional enrichment analysis was performed using clusterProfiler package. A protein-protein interaction (PPI) network of selected DEGs was constructed based on the Human Protein Reference Database. Active modules were explored using the jActiveModules plug-in in the Cytoscape Network Modeling package.In total, 537 DEGs in RA samples were identified, including 241 upregulated and 296 downregulated genes. A total of 24,451 PPI pairs were collected, and 5 active modules were screened. Furthermore, 19 submodules were acquired from the 5 active modules. Discs large homolog 1 (DLG1) and related DEGs such as guanylate cyclase 1, soluble, alpha 2 (GUCY1A2), N-methyl d-aspartate receptor 2A subunit (GRIN2A), and potassium voltage-gated channel member 1 (KCNA1) were identified in 8 submodules. Plasminogen (PLG) and related DEGs such as chemokine (C-X-C motif) ligand 2 (CXCL2), laminin, alpha 3 (LAMA3), complement component 7 (C7), and coagulation factor X (F10) were identified in 4 submodules.Our results indicate that DLG1, GUCY1A2, GRIN2A, KCNA1, PLG, CXCL2, LAMA3, C7, and F10 may play key roles in the progression of RA and may serve as putative therapeutic targets for treating RA.